Genetic manipulation of hematopoietic stem cells has the potential to treat a variety of human diseases. However, gene transfer into stem cells of large animal models and humans has thus far proven to be difficult and inefficient. By gaining insights into the biology of the hematopoietic stem cell, this project aims to improving the efficiency of gene transfer/correction of this rare and elusive target cell. Potential applications of this technology include therapeutic approaches for common blood disorders (e.g. thalassemia), malignancies (e.g. leukemia) and infectious diseases of the blood cells, such as HIV-1 infection/AIDS. We have established in vitro and in vivo animal models of human hematopoiesis using human hematopoietic progenitors obtained from cord blood or bone marrow of volunteer donors. These cells are cultured and genetically engineered in vitro, and then allowed to differentiate into mature cell lineages in vitro (using colony forming unit or T cell differentiation assays) or in vivo, using mouse or sheep animal models. These experiments allow testing vectors and gene transfer conditions that efficiently target human hematopoietic cells and determination of the capability of engineered cells to engraft and/or differentiate into mature progenies.